Gut microbiota modulation enhances the immune capacity of lizards under climate warming

Background Host-microbial interactions are expected to affect species’ adaptability to climate change but have rarely been explored in ectothermic animals. Some studies have shown that short-term warming reduced gut microbial diversity that could hamper host functional performance. Results However, our longitudinal experiments in semi-natural conditions demonstrated that warming decreased gut microbiota diversity at 2 months, but increased diversity at 13 and 27 months in a desert lizard (Eremias multiocellata). Simultaneously, long-term warming significantly increased the antibacterial activity of serum, immune responses (higher expression of intestinal immune-related genes), and the concentration of short-chain fatty acids (thereby intestinal barrier and immunity) in the lizard. Fecal microbiota transplant experiments further revealed that increased diversity of gut microbiota significantly enhanced antibacterial activity and the immune response of lizards. More specifically, the enhanced immunity is likely due to the higher relative abundance of Bacteroides in warming lizards, given that the bacteria of Bacteroides fragilis regulated IFN-β expression to increase the immune response of lizards under a warming climate. Conclusions Our study suggests that gut microbiota can help ectotherms cope with climate warming by enhancing host immune response, and highlights the importance of long-term studies on host-microbial interactions and their biological impacts. Graphical Abstract Video Abstract Supplementary Information The online version contains supplementary material available at 10.1186/s40168-023-01736-2.


Alpha diversity of gut microbiota in lizards over three periods of climate warming
To illustrate the stability of our results, we calculated chao1, Faith_PD, and Shannon indexes to characterize the alpha diversity of gut microbiota in lizards (Table S3).We used chao1 to measure community richness and found a significant decrease after 2 months in the warming climate treatment (P = 0.010, Fig. S2d), but a significant increase in community richness after 13 months (P = 0.025, Fig. S2e) and 27 months (P = 0.001, Fig. S2f) in the warming climate compared to the present climate group.We used Faith_PD to measure phylogenetic diversity and found a significant decrease after 2 months (P = 0.016, Fig. S2g), a significant increase after 27 months (P = 0.001, Fig. S2i), but no significant difference after 13 months (P = 0.201, Fig. S2h) in the warming climate group compared to the present climate group.The Shannon index was used to measure community diversity and we found a significant increase after 27 months (P = 0.002, Fig. S2l), but no significant difference after 2 months (P = 0.748, Fig. S2j) or 13 months (P = 0.261, Fig. S2k) in the warming climate group compared to the present climate group.Due to the unequal number of females and males in the climate treatments, we examined the effects of gender on the gut microbiota of lizards from the same climate.NMDS analysis showed that there was no significant difference in the composition of gut microbiota between genders in the present (P = 0.398, stress = 0.110) or warming climate (P = 0.332, stress = 0.110) treatments.

Biomarker and functional prediction of gut microbiota in lizards
In order to determine which functional pathways of lizard gut bacterial communities were involved in warming climate adaptation, we performed KEGG metabolic pathway prediction analysis.
Compared to lizards in the 2-month present climate treatment, the warming climate izards significantly up-regulated seven metabolic pathways, including cyanoamino acid metabolism, staphylococcus aureus infection, and chlorocyclohexane and chlorobenzene degradation pathways (Fig. S3a and Table S5).Lizards from the 13-month warming climate group significantly upregulated five metabolic pathways, including endocytosis, ECM−receptor interaction, and nitrotoluene degradation (Fig. S3c and Table S5).
To screen for biomarkers in lizard gut microbiota, we conducted LEfSe analyses and time series analysis (Table S4).LDA score > 3.2 showed that warming influenced lizard gut microbiota, from phylum to genus.Taxa from the phylum Firmicutes, class Clostridia, genus Roseburia, and genus S4a and b).The relative abundance of Bacteroidetes (P = 0.012, Fig. S4e and f) was significantly higher in the 27-month warming climate group than the present climate group, but the two groups did not differ in Bacteroidetes abundance after 13 months (Fig. S4c and d).
At the generic level, the relative abundance of Desulfovibrio (P = 0.012) and Odoribacter (P = 0.040) were significantly decreased in the 2-month warming climate, but the abundance of Roseburia (P = 0.026) was significantly increased (Fig. S5a and b).The relative abundances of Bacteroides (P = 0.010), Eisenbergiella (P = 0.012) and Parabacteroides (P = 0.001) were significantly higher in the 27-month warming climate group than the present climate group of lizards (Fig. S5e and f).The relative abundance of Clostridium innocuum was significantly decreased in the 13-month warming climate lizards (P = 0.031, Fig. S5c and d).(g-i) The alpha diversity index (Faith_PD) of lizard gut microbiota at 2 months (g), 13 months (h), and 27 months (i).

Fig. S2 .
Fig. S2.Effects of climate warming on environmental microbiota and gut microbiota of lizards.EP represent environment of present climate.EW represent environment of warming climate.P represents present climate.W represents warming climate.The significance of warming effects was evaluated by a two-sided

Fig. S3 .
Fig. S3.Differential microbiota and KEGG pathways between two climate groups over 2 months and 13 months.P represents present climate.W represents warming climate.(a, c) The functional profiles of gut microbiota identified by PICRUSt2 using KEGG database in different climates at 2 months (a) and 13 months (c).(b, d) Biomarkers of discriminative bacteria in different climates identified by LEfSe analysis (LDA score ≥3.2) at 2 months (b) and 13 months (d).(e) The score is plotted as A (1 -[absolute correlation coefficient]) versus B (integrated difference between the curves).Important taxonomic groups are marked with the taxa name in black.The color indicates the density of data points.(f) Temporal variation of specific bacteria communities.Y-axis represents the log2 (Fold change) of the warming climate group relative to the present climate group.X-axis represents the sampling time.The curve is simulated by loess with span as 0.8.

Fig. S6 .
Fig. S6.Survival curves of lizards exposed to two climate treatments.The solid blue line represents the median survival of lizards in the present climate group, and the yellow dashed line represents the median survival of lizards in the warming climate group.Shadows represent 95% confidence intervals.The significance of warming effects was evaluated by generalized linear mixed models with binomial distributions.